# this cell is tagged as parameters for `papermill` parameterization
# input configs
altair_config = None
nipah_config = None

# input files
entropy_file = None
func_scores_E2_file = None
binding_E2_file = None
func_scores_E3_file = None
binding_E3_file = None

# output files
filtered_E2_binding_data = None
filtered_E3_binding_data = None
filtered_E2_binding_low_effect = None
filtered_E3_binding_low_effect = None

# output images
entry_binding_combined_corr_plot = None
entry_binding_combined_corr_plot_agg = None
E2_E3_correlation = None
E2_E3_correlation_site = None
combined_E2_E3_site_corr = None
binding_by_site_plot = None
entry_binding_corr_heatmap = None
binding_corr_heatmap = None
binding_region_boxplot_plot = None
binding_region_bubble_plot = None
max_binding_in_stalk = None
max_binding_in_contact = None

# Parameters
nipah_config = "nipah_config.yaml"
altair_config = "data/custom_analyses_data/theme.py"
entropy_file = "results/entropy/entropy.csv"
func_scores_E2_file = "results/func_effects/averages/CHO_EFNB2_low_func_effects.csv"
binding_E2_file = "results/receptor_affinity/averages/EFNB2_monomeric_mut_effect.csv"
func_scores_E3_file = "results/func_effects/averages/CHO_EFNB3_low_func_effects.csv"
binding_E3_file = "results/receptor_affinity/averages/EFNB3_dimeric_mut_effect.csv"
filtered_E2_binding_data = "results/filtered_data/E2_binding_filtered.csv"
filtered_E3_binding_data = "results/filtered_data/E3_binding_filtered.csv"
filtered_E2_binding_low_effect = (
    "results/filtered_data/E2_binding_low_effect_filter.csv"
)
filtered_E3_binding_low_effect = (
    "results/filtered_data/E3_binding_low_effect_filter.csv"
)
entry_binding_combined_corr_plot = (
    "results/images/entry_binding_combined_corr_plot.html"
)
entry_binding_combined_corr_plot_agg = (
    "results/images/entry_binding_combined_corr_plot_agg.html"
)
E2_E3_correlation = "results/images/E2_E3_correlation.html"
E2_E3_correlation_site = "results/images/E2_E3_correlation_site.html"
combined_E2_E3_site_corr = "results/images/combined_E2_E3_site_corr.html"
binding_by_site_plot = "results/images/binding_by_site_plot.html"
entry_binding_corr_heatmap = "results/images/entry_binding_corr_heatmap.html"
binding_corr_heatmap = "results/images/binding_corr_heatmap.html"
binding_region_boxplot_plot = "results/images/binding_region_boxplot_plot.html"
binding_region_bubble_plot = "results/images/binding_region_bubble_plot.html"
max_binding_in_contact = "results/images/max_binding_in_contact.html"
max_binding_in_stalk = "results/images/max_binding_in_stalk.html"

import math
import os
import re
import altair as alt
import numpy as np
import pandas as pd
import scipy.stats
import yaml

# allow more rows for Altair
_ = alt.data_transformers.disable_max_rows()

if (
    os.getcwd()
    == "/fh/fast/bloom_j/computational_notebooks/blarsen/2023/Nipah_Malaysia_RBP_DMS/"
):
    pass
    print("Already in correct directory")
else:
    os.chdir(
        "/fh/fast/bloom_j/computational_notebooks/blarsen/2023/Nipah_Malaysia_RBP_DMS/"
    )
    print("Setup in correct directory")

Setup in correct directory

if nipah_config is None:
    ##hard paths in case don't want to run with snakemake
    print("loading hard paths")
    altair_config = "data/custom_analyses_data/theme.py"
    nipah_config = "nipah_config.yaml"
    entropy_file = "results/entropy/entropy.csv"

    # input files
    func_scores_E2_file = "results/func_effects/averages/CHO_EFNB2_low_func_effects.csv"
    binding_E2_file = (
        "results/receptor_affinity/averages/EFNB2_monomeric_mut_effect.csv"
    )
    func_scores_E3_file = "results/func_effects/averages/CHO_EFNB3_low_func_effects.csv"
    binding_E3_file = "results/receptor_affinity/averages/EFNB3_dimeric_mut_effect.csv"

    filtered_E2_binding_data = "results/filtered_data/E2_binding_filtered.csv"
    filtered_E3_binding_data = "results/filtered_data/E3_binding_filtered.csv"
    filtered_E2_binding_low_effect = (
        "results/filtered_data/E2_binding_low_effect_filter.csv"
    )
    filtered_E3_binding_low_effect = (
        "results/filtered_data/E3_binding_low_effect_filter.csv"
    )

if altair_config:
    with open(altair_config, "r") as file:
        exec(file.read())

with open(nipah_config) as f:
    config = yaml.safe_load(f)

# import binding and entry data
e2 = pd.read_csv(binding_E2_file)
e2_func = pd.read_csv(func_scores_E2_file)
e3 = pd.read_csv(binding_E3_file)
e3_func = pd.read_csv(func_scores_E3_file)

def merge_func_binding_dfs(func, binding, name):
    df_int = pd.merge(
        binding,
        func,
        on=["site", "mutant", "wildtype"],
        suffixes=["_binding", "_cell_entry"],
        validate="one_to_one",
        how="outer",
    ).round(3)
    df = df_int.rename(
        columns={
            "Ephrin binding_mean": "binding_mean",
            "Ephrin binding_std": "binding_std",
            "Ephrin binding_median": "binding_median",
        }
    )

    # Only save relevant columns
    df = df[
        [
            "site",
            "wildtype",
            "mutant",
            "binding_median",
            "binding_std",
            "times_seen_binding",
            "effect",
            "effect_std",
            "times_seen_cell_entry",
            "frac_models",
        ]
    ]

    def filter_binding_data(df):
        df_filter = df[
            (df["mutant"] != "*")
            & (df["mutant"] != "-")
            & (df["site"] != 603)
            &
            # Filter cell entry parameters
            (df["effect"] >= config["min_func_effect_for_binding"])
            & (df["times_seen_cell_entry"] >= config["func_times_seen_cutoff"])
            & (df["effect_std"] <= config["func_std_cutoff"])
            &
            # Filter binding parameters
            (df["times_seen_binding"] >= config["min_times_seen_binding"])
            & (df["binding_std"] <= config["max_binding_std"])
            & (df["frac_models"] >= config["frac_models"])
        ]
        return df_filter

    df_filter = filter_binding_data(df)

    # For pulling out low effect mutants for heatmaps later. Find mutants below func effect cutoff, but still have ok times_seen and func_std.
    def store_filtered_info(df):
        df_low_filter = df[
            (df["mutant"] != "*")
            & (df["mutant"] != "-")
            & (df["site"] != 603)
            & (df["effect"] < config["min_func_effect_for_binding"])
            & (df["times_seen_cell_entry"] >= config["func_times_seen_cutoff"])
            & (df["effect_std"] <= config["func_std_cutoff"])
        ]
        return df_low_filter

    df_low_effect_filter = store_filtered_info(df)

    # Save the filtered data to .csv
    if name == "EFNB2":
        print(name)
        df_filter.to_csv(filtered_E2_binding_data, index=False)
        df_low_effect_filter.to_csv(filtered_E2_binding_low_effect, index=False)
    else:
        df_filter.to_csv(filtered_E3_binding_data, index=False)
        df_low_effect_filter.to_csv(filtered_E3_binding_low_effect, index=False)

    return df_filter, df_low_effect_filter


# Call filtering function
df_E2_filter, df_E2_filter_missing = merge_func_binding_dfs(e2_func, e2, "EFNB2")
df_E3_filter, df_E3_filter_missing = merge_func_binding_dfs(e3_func, e3, "EFNB3")

# Now that they are filtered, merge EFNB2 and EFNB3
df_binding_effect_merge = pd.merge(
    df_E2_filter,
    df_E3_filter,
    on=["site", "wildtype", "mutant"],
    suffixes=["_E2", "_E3"],
    how="outer",
)

# display stats
display(df_binding_effect_merge.describe().round(3))

# Make a concat df of E2/E3 data for plotting later
df_E2_filter["selection"] = "EFNB2"
df_E3_filter["selection"] = "EFNB3"
df_binding_effect_concat = pd.concat([df_E2_filter, df_E3_filter])

EFNB2

# What are the top 5 highest and lowest binding mutants for EFNB2 and EFNB3?
def find_highest_lowest(df, name):
    print(f"We are analyzing {name}\n")
    tmp_df = df.sort_values(by="binding_median")
    print("These are the lowest binding mutants detected:")
    display(tmp_df.head(5))

    tmp_df_high = df.sort_values(by="binding_median", ascending=False)
    print("\nThese are the highest binding mutants detected:\n")
    display(tmp_df_high.head(10))

    # What about mutants with positive entry scores?
    tmp_df = df[df["effect"] > 0].sort_values(by="binding_median")
    print("These are the lowest binding mutants detected with positive entry scores:")
    display(tmp_df.head(5))

    tmp_df_high = df[df["effect"] > 0].sort_values(by="binding_median", ascending=False)
    print(
        "\nThese are the highest binding mutants detected with positive entry scores:\n"
    )
    display(tmp_df_high.head(10))


find_highest_lowest(df_E2_filter, "EFNB2")
find_highest_lowest(df_E3_filter, "EFNB3")

We are analyzing EFNB2

These are the lowest binding mutants detected:

These are the highest binding mutants detected:

These are the lowest binding mutants detected with positive entry scores:

These are the highest binding mutants detected with positive entry scores:

We are analyzing EFNB3

These are the lowest binding mutants detected:

These are the highest binding mutants detected:

These are the lowest binding mutants detected with positive entry scores:

These are the highest binding mutants detected with positive entry scores:

# Compare E2 and E3 binders
def find_highest_lowest(df):
    df["binding_diff"] = (df["binding_median_E2"] - df["binding_median_E3"]).abs()
    print(
        "These are the mutants with the biggest difference between EFNB2 and EFNB3:\n"
    )
    display(df.sort_values(by="binding_diff", ascending=False).head(5))

    # calculate aggregate differences
    agg_df = (
        df.groupby("site")[["binding_median_E2", "binding_median_E3", "binding_diff"]]
        .max()
        .reset_index()
    )
    print("These are the sites with the biggest difference between EFNB2 and EFNB3:\n")
    display(agg_df.sort_values(by="binding_diff", ascending=False).head(5))


find_highest_lowest(df_binding_effect_merge)
# find_highest_lowest(df_E3_filter,'EFNB3')

These are the mutants with the biggest difference between EFNB2 and EFNB3:

These are the sites with the biggest difference between EFNB2 and EFNB3:

def plot_corr_binding_entry_updated(df, flag):
    variant_selector = alt.selection_point(
        on="mouseover", empty=False, fields=["site", "mutant"], value=0
    )
    variant_selector_agg = alt.selection_point(
        on="mouseover", empty=False, fields=["site"], value=0
    )
    slider = alt.binding_range(min=2, max=10, step=1, name="times seen")
    selector = alt.param(name="SelectorName", value=2, bind=slider)

    empty_chart = []
    for cell in list(df["selection"].unique()):
        tmp_df = df[df["selection"] == cell]
        if flag == True:
            agg_df = (
                tmp_df.groupby("site")[["binding_median", "effect"]].sum().reset_index()
            )
            chart = (
                alt.Chart(agg_df)
                .mark_point(stroke="black", filled=True)
                .encode(
                    x=alt.X(
                        "effect",
                        title=f"Summed {cell} Cell Entry",
                        axis=alt.Axis(grid=True),
                    ),
                    y=alt.Y(
                        "binding_median",
                        title=f"Summed {cell} Binding",
                        axis=alt.Axis(grid=True),
                    ),
                    opacity=alt.condition(
                        variant_selector_agg, alt.value(1), alt.value(0.2)
                    ),
                    size=alt.condition(
                        variant_selector_agg, alt.value(100), alt.value(50)
                    ),
                    strokeWidth=alt.condition(
                        variant_selector_agg, alt.value(1), alt.value(0)
                    ),
                    color=alt.condition(
                        variant_selector_agg, alt.value("orange"), alt.value("black")
                    ),
                    tooltip=["site", "binding_median", "effect"],
                )
                .add_params(variant_selector_agg)
            )

            empty_chart.append(chart)

        else:
            chart = (
                alt.Chart(tmp_df)
                .mark_point(stroke="black", filled=True)
                .encode(
                    x=alt.X(
                        "effect", title=f"{cell} Cell Entry", axis=alt.Axis(grid=True)
                    ),
                    y=alt.Y(
                        "binding_median",
                        title=f"{cell} Binding",
                        axis=alt.Axis(grid=True),
                    ),
                    opacity=alt.condition(
                        variant_selector, alt.value(1), alt.value(0.1)
                    ),
                    size=alt.condition(variant_selector, alt.value(50), alt.value(20)),
                    strokeWidth=alt.condition(
                        variant_selector, alt.value(1), alt.value(0)
                    ),
                    color=alt.condition(
                        variant_selector, alt.value("orange"), alt.value("black")
                    ),
                    tooltip=[
                        "site",
                        "wildtype",
                        "mutant",
                        "binding_median",
                        "times_seen_binding",
                        "effect",
                    ],
                )
                .add_params(variant_selector)
            )
            empty_chart.append(chart)

    combined_chart = alt.hconcat(
        *empty_chart, title=alt.Title("Correlation between binding and entry")
    )
    return combined_chart


entry_binding_corr_plot = plot_corr_binding_entry_updated(
    df_binding_effect_concat, False
)
entry_binding_corr_plot.display()
if entry_binding_combined_corr_plot is not None:
    entry_binding_corr_plot.save(entry_binding_combined_corr_plot)

entry_binding_corr_plot_agg = plot_corr_binding_entry_updated(
    df_binding_effect_concat, True
)
entry_binding_corr_plot_agg.display()
if entry_binding_combined_corr_plot is not None:
    entry_binding_corr_plot_agg.save(entry_binding_combined_corr_plot_agg)

def plot_entry_binding_corr_heatmap(df):
    empty_chart = []

    for cell in list(df["selection"].unique()):
        tmp_df = df[df["selection"] == cell]
        chart = (
            alt.Chart(tmp_df, title=f"{cell}")
            .mark_rect()
            .encode(
                x=alt.X(
                    "effect", title="Cell Entry", axis=alt.Axis(values=[-2, -1, 0, 1])
                ).bin(maxbins=60),
                y=alt.Y(
                    "binding_median",
                    title="Binding",
                    axis=alt.Axis(values=[-4, -2, 0, 2]),
                ).bin(maxbins=60),
                color=alt.Color("count()", title="Count").scale(scheme="greenblue"),
            )
        )
        empty_chart.append(chart)

    combined_chart = alt.hconcat(
        *empty_chart, title=alt.Title("Correlation between binding and entry")
    ).resolve_scale(y="shared", x="shared", color="shared")
    return combined_chart


entry_binding_corr_heat = plot_entry_binding_corr_heatmap(df_binding_effect_concat)
entry_binding_corr_heat.display()
if entry_binding_combined_corr_plot is not None:
    entry_binding_corr_heat.save(entry_binding_corr_heatmap)

def overall_stats(df, effect, name):
    # Now group sites and find sites where all mutants are deleterious
    filtered_df = df.groupby("site").filter(lambda group: (group[effect] < -0.25).all())
    # Which sites are these?
    unique = filtered_df["site"].unique()
    # Convert unique to a Pandas Series
    unique_series = pd.Series(unique)

    # Find the common elements that are also contact sites
    unique_contact_bool = unique_series.isin(config["contact_sites"])
    # Filter and get the common elements
    common_elements = unique_series[unique_contact_bool]

    print(f"The dataset we are analyzing is: {name}\n")

    # Print the common elements
    print(
        f"Here are the contact sites that only have negative binding scores: {list(common_elements)}\n"
    )

    print(f"There are {len(unique)} sites with all negative binding score mutants\n")
    print(
        f"These are the sites with all negative binding score mutants: {list(unique)}\n"
    )

    # Now find sites with low and high binding (median)
    median_df = (
        df.groupby("site")["binding_median"]
        .max()
        .reset_index()
        .sort_values(by="binding_median", ascending=False)
    )
    print("These are the sites with the highest binding mutants:\n")
    display(median_df.head(5))

    # Now calculate mutant number
    total_mutants = df.shape[0]

    mutants_above_cutoff_tolerated = df[df["effect"] > 0]
    mutants_above_cutoff_tolerated = mutants_above_cutoff_tolerated[
        ["site", "effect", "binding_median", "wildtype", "mutant"]
    ]

    total_sites = df["site"].unique().shape[0]

    print(f"The total number of sites are: {total_sites}")


overall_stats(df_E2_filter, "binding_median", "EFNB2")
overall_stats(df_E3_filter, "binding_median", "EFNB3")

The dataset we are analyzing is: EFNB2

Here are the contact sites that only have negative binding scores: [238, 239, 242, 389, 488, 490, 491, 501, 504, 505, 531, 532, 533, 557, 579, 581, 588]

There are 43 sites with all negative binding score mutants

These are the sites with all negative binding score mutants: [116, 220, 236, 238, 239, 242, 243, 248, 346, 351, 352, 389, 390, 398, 399, 400, 435, 438, 441, 460, 467, 486, 487, 488, 490, 491, 494, 495, 497, 501, 504, 505, 526, 531, 532, 533, 557, 579, 581, 584, 585, 588, 590]

These are the sites with the highest binding mutants:

The total number of sites are: 510
The dataset we are analyzing is: EFNB3

Here are the contact sites that only have negative binding scores: [389, 488, 501, 531, 532]

There are 15 sites with all negative binding score mutants

These are the sites with all negative binding score mutants: [108, 140, 352, 389, 467, 486, 488, 494, 495, 497, 501, 510, 531, 532, 584]

These are the sites with the highest binding mutants:

The total number of sites are: 504

# find sites that are different
def find_biggest_differences(df):
    efnb2_good_efnb3_bad = df[
        (df["binding_median_E2"] > 0.5) & (df["binding_median_E3"] < -0.5)
    ].sort_values(by="binding_median_E2", ascending=False)
    display(efnb2_good_efnb3_bad)

    efnb2_bad_efnb3_good = df[
        (df["binding_median_E2"] < -0.5) & (df["binding_median_E3"] > 0.5)
    ].sort_values(by="binding_median_E3", ascending=False)
    display(efnb2_bad_efnb3_good)


find_biggest_differences(df_binding_effect_merge)

def plot_entry_binding_corr(df):
    chart = (
        alt.Chart(df, title="Correlation Between Mutant Binding Scores")
        .mark_rect()
        .encode(
            x=alt.X(
                "binding_median_E2",
                title="EFNB2 binding",
                axis=alt.Axis(values=[-5, 0, 2]),
            ).bin(maxbins=40),
            y=alt.Y(
                "binding_median_E3",
                title="EFNB3 binding",
                axis=alt.Axis(values=[-2, 0, 2]),
            ).bin(maxbins=40),
            color=alt.Color("count()", title="Count").scale(scheme="greenblue"),
        )
    )
    return chart


entry_binding_corr_heatmap_1 = plot_entry_binding_corr(df_binding_effect_merge)
entry_binding_corr_heatmap_1.display()
if entry_binding_combined_corr_plot is not None:
    entry_binding_corr_heatmap_1.save(binding_corr_heatmap)

def plot_affinity_solid(df):
    df = df.dropna()
    # calculate r value
    slope, intercept, r_value, p_value, std_err = scipy.stats.linregress(
        df["binding_median_E2"], df["binding_median_E3"]
    )
    r_value = float(r_value)
    # make chart
    chart = (
        alt.Chart(
            df,
            title=alt.Title(
                "Correlation between Mutant Binding Scores", subtitle=f"r={r_value:.2f}"
            ),
        )
        .mark_point(color="black", size=30, opacity=0.2, filled=True)
        .encode(
            x=alt.X("binding_median_E2", title=("EFNB2 Binding")),
            y=alt.Y("binding_median_E3", title=("EFNB3 Binding")),
            tooltip=[
                "site",
                "wildtype",
                "mutant",
                "binding_median_E2",
                "binding_median_E3",
                "effect_E2",
                "effect_E3",
            ],
        )
    )
    min = int(df["binding_median_E2"].min())
    max = int(df["binding_median_E3"].max())
    text = (
        alt.Chart({"values": [{"x": min, "y": max, "text": f"r = {r_value:.2f}"}]})
        .mark_text(align="left", baseline="top", dx=-10, dy=-20)
        .encode(x=alt.X("x:Q"), y=alt.Y("y:Q"), text="text:N")
    )
    chart_and_text = chart
    return chart_and_text


E2_E3_corr = plot_affinity_solid(df_binding_effect_merge)
E2_E3_corr.display()
if entry_binding_combined_corr_plot is not None:
    E2_E3_corr.save(E2_E3_correlation)

def plot_affinity_solid_mean(df):
    df = df.dropna()
    means = (
        df.groupby("site")
        .agg(
            {
                "effect_E2": "median",
                "effect_E3": "median",
                "binding_median_E2": "median",
                "binding_median_E3": "median",
                "wildtype": "first",
            }
        )
        .reset_index()
    )
    slope, intercept, r_value, p_value, std_err = scipy.stats.linregress(
        means["binding_median_E2"], means["binding_median_E3"]
    )
    r_value = float(r_value)
    chart = (
        alt.Chart(
            means,
            title=alt.Title(
                "Correlation between Aggregate Mutant Binding Scores",
                subtitle=f"r={r_value:.2f}",
            ),
        )
        .mark_point(size=50, opacity=0.3)
        .encode(
            x=alt.X(
                "binding_median_E2",
                title=("Median EFNB2 Binding"),
                axis=alt.Axis(tickCount=3),
            ),
            y=alt.Y(
                "binding_median_E3",
                title=("Median EFNB3 Binding"),
                axis=alt.Axis(tickCount=3),
            ),
            tooltip=[
                "site",
                "wildtype",
                "binding_median_E2",
                "binding_median_E3",
                "effect_E2",
                "effect_E3",
            ],
        )
    )
    text = (
        alt.Chart({"values": [{"x": -3.5, "y": 0.5, "text": f"r = {r_value:.2f}"}]})
        .mark_text(align="left", baseline="top", dx=0, dy=-10)
        .encode(x=alt.X("x:Q"), y=alt.Y("y:Q"), text="text:N")
    )
    chart_and_text = chart
    return chart_and_text


E2_E3_site_corr = plot_affinity_solid_mean(df_binding_effect_merge)
E2_E3_site_corr.display()
if entry_binding_combined_corr_plot is not None:
    E2_E3_site_corr.save(E2_E3_correlation_site)

if entry_binding_combined_corr_plot is not None:
    (E2_E3_site_corr | E2_E3_corr).save(combined_E2_E3_site_corr)

def plot_affinity_by_site_median(df):
    variant_selector = alt.selection_point(
        on="mouseover", nearest=True, empty=False, fields=["site"], value=0
    )
    empty_charts = []
    for selection in ["binding_median_E2", "binding_median_E3"]:
        if selection == "binding_median_E2":
            name = "EFNB2 Binding"
        else:
            name = "EFNB3 Binding"
        mean = df.groupby("site")[selection].max().reset_index()
        mean = mean[mean[selection] >= 0]
        chart = (
            alt.Chart(mean)
            .mark_point(stroke="black", filled=True, size=50)
            .encode(
                x=alt.X(
                    "site",
                    title=("Site"),
                    axis=alt.Axis(grid=True, tickCount=4),
                    scale=alt.Scale(domain=[70, 602]),
                ),
                y=alt.Y(selection, title=(name), axis=alt.Axis(grid=True, tickCount=3)),
                tooltip=["site"],
                color=alt.condition(
                    variant_selector, alt.value("orange"), alt.value("black")
                ),
                opacity=alt.condition(variant_selector, alt.value(1), alt.value(0.5)),
                strokeWidth=alt.condition(variant_selector, alt.value(1), alt.value(0)),
            )
            .properties(height=150, width=500)
            .add_params(variant_selector)
        )
        empty_charts.append(chart)
    combined_chart = alt.vconcat(*empty_charts, spacing=1, title="Max Binding by Site")
    return combined_chart


binding_by_site = plot_affinity_by_site_median(df_binding_effect_merge)
binding_by_site.display()
if entry_binding_combined_corr_plot is not None:
    binding_by_site.save(binding_by_site_plot)

def plot_affinity_by_contact_site(df, sites_to_show, title_text):
    variant_selector = alt.selection_point(
        on="mouseover", nearest=True, empty=False, fields=["site"], value=0
    )
    empty_charts = []

    contact_df = df[df["site"].isin(sites_to_show)]
    sites = list(contact_df["site"].unique())

    for selection in df["selection"].unique():
        tmp_df = contact_df[contact_df["selection"] == selection]
        mean = tmp_df.groupby("site")["binding_median"].max().reset_index()

        chart = (
            alt.Chart(mean)
            .mark_point(size=100)
            .encode(
                x=alt.X(
                    "site:O",
                    sort=sites,
                    title=("Site"),
                    axis=alt.Axis(grid=True, labelAngle=-90),
                    scale=alt.Scale(domain=sites),
                ),
                y=alt.Y(
                    "binding_median", title=(f"{selection}"), axis=alt.Axis(grid=True)
                ),
                tooltip=["site"],
                color=alt.condition(
                    variant_selector, alt.value("orange"), alt.value("black")
                ),
                strokeWidth=alt.condition(variant_selector, alt.value(2), alt.value(0)),
            )
            .add_params(variant_selector)
        )
        empty_charts.append(chart)
    combined_chart = alt.vconcat(*empty_charts, spacing=1, title=title_text)
    return combined_chart


contact_binding_by_site = plot_affinity_by_contact_site(
    df_binding_effect_concat, config["contact_sites"], "Max Binding in Contact"
)
contact_binding_by_site.display()
if entry_binding_combined_corr_plot is not None:
    contact_binding_by_site.save(max_binding_in_contact)

contact_binding_by_site_stalk = plot_affinity_by_contact_site(
    df_binding_effect_concat, list(range(96, 147)), "Max Binding in Stalk"
)
contact_binding_by_site_stalk.display()
if entry_binding_combined_corr_plot is not None:
    contact_binding_by_site_stalk.save(max_binding_in_stalk)

def make_boxplot_binding_region(
    df, title
):  # Create a box plot using Altair for aggregated means
    barrel_ranges = {
        "Hydrophobic": config["hydrophobic"],
        "Salt Bridges": config["salt_bridges"],
        "Hydrogen Bonds": config["h_bond_total"],
        "Contact": config["contact_sites"],
        "Overall": list(range(71, 602)),
    }

    mean_df = df.groupby("site")[["binding_median"]].median().reset_index()
    custom_order = [
        "Hydrophobic",
        "Salt Bridges",
        "Hydrogen Bonds",
        "Contact",
        "Overall",
    ]
    agg_means = []

    # For each barrel, filter the site_means dataframe to the sites belonging to that barrel and then store the means
    for barrel, sites in barrel_ranges.items():
        subset = mean_df[mean_df["site"].isin(sites)]
        for _, row in subset.iterrows():
            agg_means.append(
                {"barrel": barrel, "effect": row["binding_median"], "site": row["site"]}
            )
        agg_means_df = pd.DataFrame(agg_means)
    chart = (
        alt.Chart(agg_means_df)
        .mark_point(size=50, opacity=0.4)
        .encode(
            x=alt.X(
                "barrel:O", sort=custom_order, title=None, axis=alt.Axis(labelAngle=-90)
            ),
            y=alt.Y(
                "effect",
                title=f"Median {title} Binding",
                axis=alt.Axis(grid=True, tickCount=4),
            ),
            xOffset="random:Q",
            tooltip=["barrel", "effect", "site"],
        )
        .transform_calculate(random="sqrt(-1*log(random()))*cos(2*PI*random())")
    )

    return chart.display()


make_boxplot_binding_region(df_E2_filter, "EFNB2")
make_boxplot_binding_region(df_E3_filter, "EFNB3")

def make_boxplot_binding_region(df):
    barrel_ranges = {
        "Stalk": list(range(96, 147)),
        "Neck": list(range(148, 165)),
        "Linker": list(range(166, 177)),
        "Head": list(range(178, 602)),
        "Receptor Contact": config["contact_sites"],
        "Total": list(range(71, 602)),
    }
    custom_order = ["Stalk", "Neck", "Linker", "Head", "Receptor Contact", "Total"]
    empty_charts = []
    for selection in df["selection"].unique():
        tmp_df = df[df["selection"] == selection]
        agg_means = []

        # For each barrel, filter the site_means dataframe to the sites belonging to that barrel and then store the means
        for barrel, sites in barrel_ranges.items():
            subset = tmp_df[tmp_df["site"].isin(sites)]
            for _, row in subset.iterrows():
                agg_means.append(
                    {
                        "region": barrel,
                        "binding_median": row["binding_median"],
                        "site": row["site"],
                    }
                )
            agg_means_df = pd.DataFrame(agg_means)

        chart = (
            alt.Chart(agg_means_df, title=f"{selection}")
            .mark_boxplot(color="darkgray", extent="min-max", opacity=1)
            .encode(
                x=alt.X(
                    "region:O",
                    sort=custom_order,
                    title="RBP Region",
                    axis=alt.Axis(labelAngle=-90),
                ),
                y=alt.Y(
                    "binding_median",
                    title=f"Binding",
                    axis=alt.Axis(grid=True, tickCount=4),
                ),
                tooltip=["region", "binding_median", "site"],
            )
            .properties(width=config["width"], height=config["height"])
        )
        empty_charts.append(chart)
    combined_effect_chart = alt.hconcat(*empty_charts).resolve_scale(
        y="shared", x="shared", color="independent"
    )
    return combined_effect_chart


entry_region_boxplot = make_boxplot_binding_region(df_binding_effect_concat)
entry_region_boxplot.display()
if entry_binding_combined_corr_plot is not None:
    entry_region_boxplot.save(binding_region_boxplot_plot)

def make_bubble_binding_region(df):
    barrel_ranges = {
        "Stalk": list(range(96, 147)),
        "Neck": list(range(148, 165)),
        "Linker": list(range(166, 177)),
        "Head": list(range(178, 602)),
        "Receptor Contact": config["contact_sites"],
        "Total": list(range(71, 602)),
    }
    custom_order = ["Stalk", "Neck", "Linker", "Head", "Receptor Contact", "Total"]
    empty_charts = []
    for selection in df["selection"].unique():
        tmp_df = df[df["selection"] == selection]
        agg_means = []

        # For each barrel, filter the site_means dataframe to the sites belonging to that barrel and then store the means
        for barrel, sites in barrel_ranges.items():
            subset = tmp_df[tmp_df["site"].isin(sites)]
            for _, row in subset.iterrows():
                agg_means.append(
                    {
                        "region": barrel,
                        "binding_median": row["binding_median"],
                        "site": row["site"],
                        "mutant": row["mutant"],
                    }
                )
            agg_means_df = pd.DataFrame(agg_means)

        variant_selector = alt.selection_point(
            on="mouseover", empty=False, fields=["site", "mutant"], value=1
        )

        chart = (
            alt.Chart(agg_means_df, title=f"{selection}")
            .mark_point(opacity=0.3, stroke="black")
            .encode(
                x=alt.X(
                    "region:O",
                    sort=custom_order,
                    title="RBP Region",
                    axis=alt.Axis(labelAngle=-90),
                ),
                y=alt.Y(
                    "binding_median",
                    title=f"Binding",
                    axis=alt.Axis(grid=True, tickCount=4),
                ),
                xOffset="random:Q",
                tooltip=["region", "binding_median", "site", "mutant"],
                color=alt.condition(
                    variant_selector, alt.value("orange"), alt.value("black")
                ),
                opacity=alt.condition(variant_selector, alt.value(1), alt.value(0.1)),
                strokeWidth=alt.condition(variant_selector, alt.value(2), alt.value(0)),
                size=alt.condition(variant_selector, alt.value(50), alt.value(15)),
            )
            .transform_calculate(random="sqrt(-1*log(random()))*cos(2*PI*random())")
            .properties(width=config["width"], height=config["height"])
        ).add_params(variant_selector)
        empty_charts.append(chart)
    combined_effect_chart = (
        alt.hconcat(*empty_charts)
        .resolve_scale(y="shared", x="shared", color="independent")
        .add_params(variant_selector)
    )
    return combined_effect_chart


entry_region_bubble = make_bubble_binding_region(df_binding_effect_concat)
entry_region_bubble.display()
if entry_binding_combined_corr_plot is not None:
    entry_region_bubble.save(binding_region_bubble_plot)

	site	binding_median_E2	binding_std_E2	times_seen_binding_E2	effect_E2	effect_std_E2	times_seen_cell_entry_E2	frac_models_E2	binding_median_E3	binding_std_E3	times_seen_binding_E3	effect_E3	effect_std_E3	times_seen_cell_entry_E3	frac_models_E3
count	7212.000	6684.000	6684.000	6684.000	6684.000	6684.000	6684.000	6684.000	6519.000	6519.000	6519.000	6519.000	6519.000	6519.000	6519.0
mean	343.684	-0.333	0.501	6.400	-0.061	0.368	7.699	0.998	-0.017	0.179	6.227	-0.039	0.388	6.804	1.0
std	148.183	1.095	0.321	3.203	0.495	0.191	4.383	0.022	0.271	0.169	3.053	0.474	0.186	3.584	0.0
min	71.000	-5.494	0.008	2.250	-1.500	0.010	2.000	0.750	-2.147	0.000	2.500	-1.500	0.033	2.000	1.0
25%	217.000	-0.416	0.273	4.500	-0.335	0.223	5.000	1.000	-0.150	0.062	4.500	-0.266	0.248	4.714	1.0
50%	342.000	-0.033	0.426	5.750	0.076	0.337	6.750	1.000	-0.012	0.135	5.500	0.094	0.356	6.000	1.0
75%	468.000	0.190	0.645	7.500	0.329	0.480	9.125	1.000	0.118	0.241	7.500	0.324	0.496	7.857	1.0
max	602.000	2.205	1.995	43.750	0.617	0.994	72.380	1.000	2.006	1.780	45.000	0.616	1.000	56.710	1.0

	site	wildtype	mutant	binding_median	binding_std	times_seen_binding	effect	effect_std	times_seen_cell_entry	frac_models	selection
8285	501	E	K	-5.494	0.928	24.50	-0.712	0.606	32.00	1.0	EFNB2
8898	532	A	V	-5.180	1.140	16.25	-0.292	0.312	17.88	1.0	EFNB2
8072	490	Q	K	-4.871	1.081	8.50	-0.673	0.539	11.38	1.0	EFNB2
8070	490	Q	H	-4.864	0.862	8.25	-0.273	0.668	10.88	1.0	EFNB2
3188	236	R	H	-4.853	0.883	22.00	-1.217	0.335	29.38	1.0	EFNB2

	site	wildtype	mutant	binding_median	binding_std	times_seen_binding	effect	effect_std	times_seen_cell_entry	frac_models	selection
5459	354	S	R	2.205	1.165	4.25	-0.922	0.537	6.750	1.00	EFNB2
9830	580	I	S	2.165	1.768	4.75	-0.750	0.692	6.375	1.00	EFNB2
9948	586	N	T	2.096	0.578	5.25	-0.658	0.823	7.000	1.00	EFNB2
655	104	E	T	2.094	0.755	4.50	-0.348	0.671	3.500	1.00	EFNB2
2696	211	G	F	2.060	0.549	4.75	-0.753	0.426	5.125	1.00	EFNB2
2621	207	L	I	2.042	1.531	4.00	-0.816	0.764	3.750	1.00	EFNB2
1298	138	I	L	1.968	0.917	5.00	-1.114	0.244	4.750	1.00	EFNB2
2051	178	V	H	1.938	0.730	3.25	-0.639	0.903	3.625	1.00	EFNB2
9307	553	S	W	1.923	0.287	10.00	-0.396	0.429	13.250	1.00	EFNB2
4768	318	R	P	1.922	1.433	3.00	-0.961	0.764	4.000	0.75	EFNB2

	site	wildtype	mutant	binding_median	binding_std	times_seen_binding	effect	effect_std	times_seen_cell_entry	frac_models	selection
9980	588	I	L	-4.684	0.874	5.750	0.159	0.420	6.375	1.0	EFNB2
9402	558	A	V	-4.660	0.730	17.250	0.152	0.364	18.500	1.0	EFNB2
9369	557	N	D	-4.637	0.399	9.500	0.547	0.479	11.120	1.0	EFNB2
8146	494	P	E	-4.598	0.874	7.812	0.287	0.194	9.750	1.0	EFNB2
8773	526	L	H	-4.539	0.782	5.250	0.222	0.392	6.750	1.0	EFNB2

	site	wildtype	mutant	binding_median	binding_std	times_seen_binding	effect	effect_std	times_seen_cell_entry	frac_models	selection
9548	566	F	H	1.388	0.847	4.25	0.186	0.554	4.375	1.0	EFNB2
7303	450	Q	I	1.330	1.194	5.00	0.316	0.399	6.000	1.0	EFNB2
10034	591	K	G	1.266	0.156	5.75	0.027	0.442	6.125	1.0	EFNB2
4540	306	N	T	1.239	0.491	4.00	0.110	0.357	7.625	1.0	EFNB2
9513	564	N	K	1.196	0.488	7.25	0.228	0.442	8.000	1.0	EFNB2
2034	177	G	M	1.193	0.618	6.50	0.380	0.534	5.375	1.0	EFNB2
7184	444	I	F	1.162	0.350	5.00	0.294	0.314	6.250	1.0	EFNB2
2519	202	L	F	1.156	0.358	3.75	0.096	0.344	4.375	1.0	EFNB2
4594	309	Y	P	1.148	1.065	3.00	0.389	0.521	2.875	1.0	EFNB2
9764	577	L	I	1.115	0.101	5.75	0.041	0.398	5.500	1.0	EFNB2

Analyze selection data using soluble Ephrin-B2 or -B3¶

hard paths for running in interactive mode¶

Run config files to setup altair theme and config variables¶

Import the binding and entry data for EFNB2 and EFNB3¶

Filter the data and save¶

Make plots showing correlation between binding and entry for EFNB2 and EFNB3¶

Same plot as above, but slightly different format¶

Calculate some stats on binding¶

Find sites with opposite effects on binding¶

Find correlations between EFNB2 and EFNB3 binding¶

Plot correlations between summary statistics for each site¶

Make plot showing binding by site (median)¶

Make bubble plots for binding in different areas of receptor pocket¶

make boxplot of binding scores by region¶

	site	wildtype	mutant	binding_median	binding_std	times_seen_binding	effect	effect_std	times_seen_cell_entry	frac_models	selection
9221	555	D	K	-2.147	1.225	6.0	0.251	0.422	6.714	1.0	EFNB3
9227	555	D	R	-1.510	0.420	4.0	0.312	0.526	4.143	1.0	EFNB3
8739	530	Q	L	-1.447	0.477	3.5	-1.231	0.862	5.857	1.0	EFNB3
9761	583	T	R	-1.410	0.446	5.5	-0.870	0.485	6.286	1.0	EFNB3
9755	583	T	K	-1.247	0.113	5.0	-0.667	0.644	7.000	1.0	EFNB3

	site	wildtype	mutant	binding_median	binding_std	times_seen_binding	effect	effect_std	times_seen_cell_entry	frac_models	selection
9866	589	R	G	2.006	0.659	7.0	-1.197	0.576	8.286	1.0	EFNB3
9699	580	I	M	1.337	0.534	5.5	-0.873	0.531	6.714	1.0	EFNB3
3777	270	V	Q	1.329	1.028	5.0	-0.920	0.651	5.429	1.0	EFNB3
3719	267	M	Q	1.291	1.371	4.5	-0.594	0.818	5.667	1.0	EFNB3
8010	492	Q	L	1.261	0.067	5.5	0.548	0.183	5.143	1.0	EFNB3
1150	132	S	Y	1.247	0.722	5.5	-1.024	0.573	6.000	1.0	EFNB3
9691	580	I	C	1.241	0.524	4.0	-0.173	0.401	4.857	1.0	EFNB3
9871	589	R	M	1.234	0.635	4.5	-0.726	0.342	5.429	1.0	EFNB3
9566	573	W	M	1.215	1.373	4.0	-0.835	0.676	5.429	1.0	EFNB3
9853	588	I	P	1.183	0.255	5.5	-0.607	0.382	5.429	1.0	EFNB3

	site	wildtype	mutant	binding_median_E2	binding_std_E2	times_seen_binding_E2	effect_E2	effect_std_E2	times_seen_cell_entry_E2	frac_models_E2	binding_median_E3	binding_std_E3	times_seen_binding_E3	effect_E3	effect_std_E3	times_seen_cell_entry_E3	frac_models_E3	binding_diff
5729	530	Q	F	-4.114	0.466	6.25	0.550	0.202	5.375	1.0	0.084	0.062	6.0	0.408	0.274	6.286	1.0	4.198
5258	490	Q	W	-4.519	1.005	3.75	0.407	0.378	4.750	1.0	-0.586	0.323	3.0	-0.130	0.458	3.286	1.0	3.933
5259	490	Q	Y	-4.089	0.900	5.50	-1.085	0.400	6.375	1.0	-0.461	0.574	5.5	-0.280	0.797	5.714	1.0	3.628
5263	491	S	G	-4.183	0.595	6.50	0.058	0.298	8.625	1.0	-0.562	0.047	5.5	-0.387	0.608	6.286	1.0	3.621
5283	492	Q	K	-3.484	0.549	11.00	0.430	0.232	11.620	1.0	0.006	0.077	10.0	0.388	0.240	11.570	1.0	3.490

	site	binding_median_E2	binding_median_E3	binding_diff
441	530	0.628	0.405	4.198
405	490	-0.967	0.112	3.933
406	491	-0.799	0.645	3.621
407	492	0.362	1.261	3.490
162	239	-0.393	-0.041	3.418

	site	wildtype	mutant	binding_median_E2	binding_std_E2	times_seen_binding_E2	effect_E2	effect_std_E2	times_seen_cell_entry_E2	frac_models_E2	binding_median_E3	binding_std_E3	times_seen_binding_E3	effect_E3	effect_std_E3	times_seen_cell_entry_E3	frac_models_E3	binding_diff
2555	303	P	C	1.223	1.577	5.00	-0.745	0.590	5.000	1.0	-0.566	0.098	4.0	-0.527	0.594	4.857	1.0	1.789
5937	546	L	H	1.161	1.145	5.25	-1.264	0.387	6.750	1.0	-0.543	0.665	3.5	-0.674	0.544	5.143	1.0	1.704
997	178	V	T	0.796	0.726	3.75	-0.272	0.759	4.250	1.0	-0.513	0.600	3.0	-0.022	0.425	3.571	1.0	1.309
4692	447	S	E	0.767	1.742	4.25	-1.116	0.812	4.750	1.0	-0.557	0.393	4.0	-0.078	0.325	4.571	1.0	1.324
1619	223	A	M	0.736	1.538	3.25	-0.383	0.522	3.375	1.0	-0.561	0.206	3.0	-0.085	0.408	3.571	1.0	1.297
109	79	N	S	0.692	1.652	3.00	-0.368	0.496	2.375	1.0	-0.646	0.260	3.0	-0.011	0.569	3.429	1.0	1.338
90	78	D	I	0.521	0.634	5.25	-0.894	0.398	6.500	1.0	-0.566	0.755	5.0	-0.664	0.845	5.143	1.0	1.087

	site	wildtype	mutant	binding_median_E2	binding_std_E2	times_seen_binding_E2	effect_E2	effect_std_E2	times_seen_cell_entry_E2	frac_models_E2	binding_median_E3	binding_std_E3	times_seen_binding_E3	effect_E3	effect_std_E3	times_seen_cell_entry_E3	frac_models_E3	binding_diff
6512	588	I	P	-2.042	0.840	5.25	-0.276	0.300	4.750	1.0	1.183	0.255	5.5	-0.607	0.382	5.429	1.0	3.225
3107	338	R	G	-0.574	0.507	4.50	0.458	0.422	5.250	1.0	0.684	0.976	4.0	0.270	0.340	3.429	1.0	1.258
5260	491	S	A	-0.799	0.537	5.50	-0.441	0.806	6.250	1.0	0.645	0.010	6.5	0.232	0.358	6.143	1.0	1.444
5287	492	Q	R	-2.826	0.611	19.25	-0.003	0.272	23.120	1.0	0.644	0.054	20.0	0.549	0.119	20.570	1.0	3.470
4367	424	P	A	-0.502	0.276	3.50	0.306	0.322	3.625	1.0	0.592	0.229	2.5	0.329	0.188	2.857	1.0	1.094